Approximately 16% of adult Americans have been infected by herpes simplex virus type 2 (HSV-2), the main cause of genital herpes. The virus establishes a lifelong latent infection from which it periodically reactivates and returns to the periphery where it can cause recurrent disease or be shed without clinical symptoms, resulting in efficient transmission to new individuals. Transmission of HSV-2 from mother to neonate at birth results in serious morbidity. Further, HSV-2 infections have been shown to increase the risk of HIV acquisition by 2 to 4- fold. Immunization would be the most effective approach to preventing genital herpes, but prophylactic vaccines to protect against HSV-2 disease have failed in clinical trials. Clearly new strategies are needed. Therapeutic immunization has the potential to adapt and improve normal host antiviral immune responses and impact the pathological consequences associated with a latent HSV-2 infection. Consequently, the studies that we propose in this application address a significant clinical problem which is of direct relevance to this RFA. Here we will explore the use of therapeutic immunization to reduce the burden of latent virus infection, to limit reactivation from latency, or to quickly limit the repliation of reactivated virus reducing damage in the host. Guinea pigs are the only well characterized animal model which experience spontaneous reactivation events during latent HSV-2 infection resulting in recurrent virus shedding and clinical signs similar to human disease. Thus, they represent the best model to test hypotheses of immune modulation of HSV-2 latency. Our long term goal is to develop a therapeutic vaccine approach that will reduce the burden of latent HSV-2 infection and the pathogenesis associated with the latent infection. The objective of this application is to optimize delivery of a therapeutic HSV-2 vaccine and immune, imaging, and molecular virology techniques to quantify host responses and viral pathogenesis events both at the neuronal site of HSV-2 latency and in genital tissue the site of HSV-2 re-emergence. Our central hypothesis is that therapeutic immunization with a DNA vaccine can be used to impact the establishment of HSV-2 latency and will decrease recurrent genital disease as well as the magnitude and frequency of HSV-2 shedding. Aim 1 will develop powerful new techniques to quantify important cell-mediated immune events at the neuronal site of HSV-2 latency and at the genital site of re-emergence from latency. Aim 2 will utilize these newly developed methods to determine if therapeutic immunization at the onset of primary clinical disease will improve the efficacy of a candidate therapeutic HSV-2 vaccine, resulting in significantly lower latent virus load and diminished recurrent disease. Aim 3 will establish new imaging techniques to increase our understanding of the pathogenesis of HSV-2 following re-emergence from latency and to provide important quantitative algorithms to accurately measure the effect of reactivating virus on the genital epithelium. Aim 4 will develop novel vaccine constructs designed to increase cell-mediated immune responses to immunization and to more effectively interfere with HSV-2 latency and reactivation.